In this paper, we report on a general approach for the detection of a specific tumoural biomarker directly in serum. Such detection is made possible using a protein-binding peptide selected through an improved phage display technique and then conjugated to engineered microparticles (MPs). Protein biomarkers represent an unlimited source of information for non-invasive diagnostic and prognostic tests; MP-based assays are becoming largely used in manipulation of soluble biomarkers, but their direct use in serum is hampered by the complex biomolecular environment. Our technique overcomes the current limitations as it produces a selective MP-engineered with an antifouling layer-that 'captures' the relevant protein staying impervious to the background. Our system succeeds in fishing-out the human tumour necrosis factor alpha directly in serum with a high selectivity degree. Our method could have great impact in soluble protein manipulation and detection for a wide variety of diagnostic applications.
Supercritical emulsion extraction (SEE) was recently proposed for the production of biopolymer microparticles starting from oil-in-water emulsions. This technology can improve the product quality because of the fast and selective extraction of the dispersed oily phase by using supercritical carbon dioxide (SC-CO(2) ). However, until now, SEE was proposed in batch configuration, sharing with the traditional processes an intrinsically discontinuous operation and problems of batches reproducibility and process yield. In this study, by using a countercurrent packed column, the SEE process was proposed in a continuous operating mode (SEE-CM) for the production of poly-lactic-co-glycolic acid (PLGA) microparticles. The new process design takes advantage of the large contact area between the SC-CO(2) and emulsion allowing the production of PLGA microparticles with controlled and narrow size distributions in only few minutes. SEE-CM operating parameters such as pressure, temperature, and flow rate ratios were analyzed and the process efficiency in terms of recovered material and its size distribution compared with SEE (batch mode operation) and conventional evaporation technology. PLGA microparticles showed a mean particle size between 1-3 µm (depending on the droplet sizes) with a SD that was always smaller than that associated with particles produced by discontinuous processes. Single and double emulsions were successfully treated and the microparticles physico-chemical properties showed no morphological and structural differences between the SEE-CM-produced microparticles and the ones obtained by conventional evaporation technology.
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